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. 2012 Jul 18;44(9):554–561. doi: 10.3858/emm.2012.44.9.063

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Copyright © 2012 by the Korean Society for Biochemistry and Molecular Biology

This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Immunoblot analysis and immunoprecipitation of nucleolin with AS1411-cotinine/anti-cotinine antibody complexes. (A) Immunoblot analysis. Raji cell lysates were subjected to 4-12% SDS-PAGE, and proteins were transferred onto a nitrocellulose membrane. The membrane was probed with AS1411-cotinine/anti-cotinine antibody complexes (lane 1), AS1411-cotinine/control antibody complexes (lane 2), CRO26-cotinine/anti-cotinine antibody complexes (lane 3), and mouse anti-nucleolin antibody (lane 4). HRP-conjugated mouse anti-human IgG, HRP-conjugated rabbit anti-mouse IgG antibody, and SuperSignal Pico West chemiluminescent substrate were used for visualization of the bands. (B) Immunoprecipitation. Raji cell lysates were incubated with AS1411-cotinine/anti-cotinine antibody complexes (lane 1), AS1411-cotinine/control antibody complexes (lane 2), and CRO26-cotinine/anti-cotinine antibody complexes (lane 3). All lysate mixtures were incubated with immobilized protein A beads, and precipitated proteins were fractionated by SDS-PAGE and probed with mouse anti-nucleolin antibody. In lane 4, the cell lysate was directly loaded onto the gel and probed with mouse anti-nucleolin antibody.